Hexosamine biosynthesis drives hemocyanin O-GlcNAcylation to potentiate antibacterial immunity in shrimp

Abstract Post-translational modifications (PTMs) are key regulators of immune responses, however, their roles in invertebrate immunity remain poorly defined. Here, we show that Penaeus vannamei employs O-GlcNAcylation, a dynamic PTM controlled by the hexosamine biosynthesis pathway (HBP), to enhance antibacterial defense. Bacterial infection induces metabolic reprogramming in hemocytes, upregulating HBP enzymes and promoting O-GlcNAcylation of hemocyanin (PvHMC) through O-GlcNAc transferase (PvOGT). Site-specific modification of the PvHMC large subunit at Thr584 enhances its conformational stability and interaction with bacterial pathogen-associated molecular patterns, including lipopolysaccharide and peptidoglycan, thereby increasing bacterial binding, agglutination, and killing. Disruption of HBP flux or OGT activity reduces hemocyanin O-GlcNAcylation and impairs bacterial clearance, whereas inhibition of O-GlcNAcase enhances O-GlcNAcylation and antibacterial efficacy. Together, these findings identify HBP-driven O-GlcNAcylation as a metabolic-immune regulatory axis in shrimp and establish hemocyanin O-GlcNAcylation as a key mechanisms underlying effective innate antibacterial defense, with potential implications for disease control in aquaculture.